Studies on medium-chain fatty acyl-coenzyme a synthetase. Purification and properties.
1. Medium-chain fatty acyl-CoA synthetase (EC 6.2.1.2) was isolated by the method of Mahler, Wakil & Bock (1953) and the enzyme activity determined by the disappearance of CoA in the presence either of butyrate and ATP or of butyryl-AMP, as well as by ATP formation from butyryl-AMP and PP(i). 2. Preincubation of the enzyme with CoA and ATP alone or together, followed by the removal of these substrates by gel filtration, caused a marked inhibition of ATP formation, contrary to results previously obtained with palmitoyl-CoA synthetase. 3. The effect of ATP on butyryl-AMP-dependent CoA disappearance was inconsistent. Low concentrations of ATP (0.1-0.5mm) always caused inhibition, whereas higher concentrations (5-10mm) activated in some enzyme preparations and inhibited in others. 4. This inconsistency was shown to be due to the presence of two enzyme fractions. Both fractions had similar activities when assayed by the butyryl-AMP- or butyrate-plus-ATP-dependent CoA disappearance. However, fraction I was activated by ATP as measured by butyryl-AMP-dependent CoA disappearance whereas fraction II was inhibited by it. Fraction I also catalysed ATP formation from butyryl-AMP and PP(i) whereas fraction II was lacking in such activity. 5. The relationship of these observations with respect to other known mechanisms of fatty acid-activating systems is discussed.
D000227:Adenine Nucleotides; D000255:Adenosine Triphosphate / Q000096:biosynthesis; D000818:Animals; D002087:Butyrates / Q000378:metabolism; D002417:Cattle; D002850:Chromatography, Gel; D003065:Coenzyme A / Q000378:metabolism*; D003864:Depression, Chemical; D005227:Fatty Acids / Q000378:metabolism*; D066298:In Vitro Techniques; D008025:Ligases / Q000302:isolation & purification* / Q000378:metabolism*; D008099:Liver / Q000201:enzymology; D010710:Phosphates
D000227:Adenine Nucleotides; D002087:Butyrates; D005227:Fatty Acids; D010710:Phosphates; D000255:Adenosine Triphosphate; D008025:Ligases; D003065:Coenzyme A